Department of Biology, Chungnam National University, Daejeon, 34134 Republic of Korea.
Korean Research Institute of Biosciences and Biotechnology, Daejeon, 34141 Republic of Korea.
Mol Autism. 2017 Sep 29;8:50. doi: 10.1186/s13229-017-0168-2. eCollection 2017.
maps to the Down syndrome critical region at 21q22. Mutations in this kinase-encoding gene have been reported to cause microcephaly associated with either intellectual disability or autism in humans. Intellectual disability accompanied by microcephaly was recapitulated in a murine model by overexpressing which mimicked Down syndrome phenotypes. However, given embryonic lethality in homozygous knockout (KO) mice, no murine model studies could present sufficient evidence to link dysfunction with autism. To understand the molecular mechanisms underlying microcephaly and autism spectrum disorders (ASD), we established an in vivo KO model using zebrafish.
We identified a patient with a mutation in the gene using microarray analysis. Circumventing the barrier of murine model studies, we generated a KO zebrafish using transcription activator-like effector nuclease (TALEN)-mediated genome editing. For social behavioral tests, we have established a social interaction test, shoaling assay, and group behavior assay. For molecular analysis, we examined the neuronal activity in specific brain regions of KO zebrafish through in situ hybridization with various probes including and which are the molecular markers for stress response.
Microarray detected an intragenic microdeletion of in an individual with microcephaly and autism. From behavioral tests of social interaction and group behavior, KO zebrafish exhibited social impairments that reproduce human phenotypes of autism in a vertebrate animal model. Social impairment in KO zebrafish was further confirmed by molecular analysis of and expression. Transcriptional expression of and was lower than that of wild type fish in specific hypothalamic regions, suggesting that KO fish brains are less activated by social context.
In this study, we established a zebrafish model to validate a candidate gene for autism in a vertebrate animal. These results illustrate the functional deficiency of as an underlying disease mechanism for autism. We also propose simple social behavioral assays as a tool for the broader study of autism candidate genes.
该基因定位于 21q22 的唐氏综合征关键区域。该激酶编码基因突变已被报道可导致人类小头畸形伴智力障碍或自闭症。通过过表达该基因模拟唐氏综合征表型，在小鼠模型中重现了伴有小头畸形的智力障碍。然而，由于纯合敲除（KO）小鼠的胚胎致死性，没有小鼠模型研究能够提供充分的证据将 功能障碍与自闭症联系起来。为了了解小头畸形和自闭症谱系障碍（ASD）的分子机制，我们使用斑马鱼建立了一个体内 KO 模型。
我们使用微阵列分析鉴定了一位携带 基因变异的患者。为了绕过小鼠模型研究的障碍，我们使用转录激活因子样效应物核酸酶（TALEN）介导的基因组编辑生成了一个 KO 斑马鱼。对于社会行为测试，我们建立了社会互动测试、群聚测试和群体行为测试。对于分子分析，我们通过原位杂交用各种探针（包括 和 ，它们是应激反应的分子标记物）检查了 KO 斑马鱼特定脑区的神经元活性。
微阵列检测到一个个体的微缺失，该个体患有小头畸形和自闭症。从社会互动和群体行为的行为测试来看， KO 斑马鱼表现出社交障碍，在脊椎动物动物模型中再现了人类自闭症的表型。在 KO 斑马鱼中，通过 和 表达的分子分析进一步证实了社会障碍。与野生型鱼相比，特定下丘脑区域的 和 的转录表达较低，这表明 KO 鱼的大脑对外界社会环境的刺激反应较低。
在这项研究中，我们建立了一个斑马鱼模型，在脊椎动物动物中验证了自闭症的一个候选基因。这些结果说明了 作为自闭症潜在疾病机制的功能缺陷。我们还提出了简单的社会行为测试作为自闭症候选基因更广泛研究的工具。
